The normal gastrointestinal motility depends on a cascade of intracellular signals that translate a signal at the cell membrane into muscle contraction. The calcium ion plays a central role in this process of excitation-contraction coupling. Changes in the calcium homeostasis of smooth muscle cells may thus contribute to common gastrointestinal diseases, such as constipation, diarrhea, or gastroesophageal reflux disease. Increases in the cytosolic calcium level can be due to calcium influx through ion channels in the cell membrane or calcium release from intracellular stores. Two intracellular calcium release channels have been identified in intestinal smooth muscle cells: the inositol 1,4,5-trisphosphate receptor channel and the ryanodine receptor channel. Little is known about the mechanisms that modulate the activity of these intracellular calcium release channels. We hypothesize that ryanodine receptor channel isoforms are differentially expressed in gastro-intestinal smooth muscle from anatomically and functionally distinct areas; associated modulatory proteins further increase this heterogeneity, thereby affecting intestinal motor function. The proposed experiments will investigate the regulation of calcium release from intracellular stores. The following specific goals will be addressed: (1) Biochemical and functional characterization of calcium release channels expressed in intestinal smooth muscle cells. (2) Identification of proteins associated with ryanodine receptor channels in intestinal smooth muscle cells. (3) Characterization of the functional role of proteins associated with the ryanodine receptor channel. A better understanding of mechanisms that control the calcium homeostasis may provide important insight into the etiology of diseases or lead to the development of novel treatment strategies for functional abnormalities.